Another Step Toward DIYStemCells

(18 June 2009: Lightly edited for clarity.)

The June 5 issue of Cell Stem Cells has a brief report describing the use of four proteins to reprogram human fibroblasts into induced pluripotent stem cells (iPSCs).  I think this is a pretty important paper, as it dispenses with any sort of genetic manipulation of the target cells or any use of plasmids to insert new "control circuitry", or any chemical manipulation whatsoever.

As expected, it is getting easier to produce iPSCs, and the authors of the paper ("Generation of Human Induced Pluripotent Stem Cells by Direct Delivery of Reprogramming Proteins") note that their work demonstrates the elimination of "the potential risks associated with the use of viruses, DNA transfection, and potentially harmful chemicals and in the future could potentially provide a safe source of patient-specific cells for regenerative medicine".

Kim et al used four recombinant human proteins to turn human newborn fibroblast cells (purchased from ATCC -- see the Supplemental Data) into iPSCs, where each of the proteins was fused to a nine amino acid long "cell-penetrating peptide" (CPP) that facilitated the importation of the proteins across the cell membrane.  The procedure was not particularly efficient, but after multiple treatments the authors produced cells that could differentiate into many different kinds of human tissues.

Here are a couple of thoughts about the paper.  Note that in what follows I have only had a few sips of my first cup of coffee today, and my brain is still quite fuzzy, but I think I am mostly coherent.  You can be the judge.

First, the authors did not use mature cells from adults, so don't expect this paper to lead to replacement organs and tissues tomorrow.  The use of cells from newborns makes a great deal of sense for a first go at getting protein-based reprogramming to work, as those cells have already been demonstrated to be relatively easy to reprogram.  The published procedure required many weeks of effort to produce iPSCs, and authors note that they have quite a ways to go before they can produce stem cells at the same efficiency as other techniques.

Nonetheless, it works.

Second, the paper describes PCR-based cloning of human genes to add the CPP sequences, along with a fair amount of bench manipulation to generate cells that made each of the four reprogramming proteins.  All the sequences for those proteins are online, as are the sequences for the CPPs, so generating the corresponding genes by synthesis rather than cloning would now cost less than $10K, with delivery in 2-4 weeks.  In another year, it will probably cost no more than $5K.  (How long will it be before these proteins show up in the Registry of Standard Biology Parts?)

Third, the authors did not use purified reprogramming proteins to generate iPSCs, but rather used whole cell extracts from cells that produced those proteins.  Thus the concentrations of the reprogramming proteins were limited to whatever was in the cell extract.  This might critically affect the efficiency of the reprogramming.  Presumably, the authors are already working on generating cultured cell lines to produced the reprogramming proteins in larger quantities.  But if you wanted to do it yourself, it looks like you might "simply" have to order the appropriate sequences from Blue Heron already cloned into the human expression plasmid pCDNA3.1/myc-His A, which is available from Invitrogen.  This would add a couple of hundred dollars to the cost because Blue Heron would have to play around with a proprietary plasmid instead of the public domain plasmids they usually use to ship genes.  You would then follow the recipe from the Supplementary Data to transform a protein production cell line to make those proteins.  Or perhaps you have a favorite recipe of your own.  Here is something I don't get -- it looks like that particular expression plasmid adds a His tag to the end of the gene, so I don't understand why Kim et al didn't try a purification step, but maybe that is underway.

Fourth, if you wanted to do this at home, you could.  You should expect to fail many times.  And then you should expect to fail some more.  And then, assuming your human cell culture technique is up to snuff, you should expect to eventually succeed.  You might want to wait until the inevitable paper showing how to do this with adult differentiated skin cells is published.

And then what?

You will have an autologous stem cell line that you can use to produce tissues that are, immunologically speaking, identical to those in your body.  What should you do with them?  I would suggest you show them off at cocktail parties, brag about them on Facebook, and then destroy them with bleach and an autoclave.  In lieu of an autoclave a microwave would probably do just fine.

But I expect that at least some of you will try to follow a recipe to generate some sort of human tissue, or even to simply inject those cells in your own bodies, which will result in all kinds of crazy teratomas and other tumors.  To quote Harold Ramus, "that would be bad".  So don't do that.  Just because DIYStemCells are cool doesn't mean you should actually use them yourself.  But I know some of you will anyway.  That is the future of biological technologies, for better or worse.